Altered T-cell subset distribution in the viral reservoir in HIV-1-infected individuals with extremely low proviral DNA (LoViReTs)

HIV cure strategies aim to eliminate viral reservoirs that persist despite successful antiretroviral therapy (ART). We have previously described that 9% of HIV-infected individuals who receive ART harbor low levels of provirus (LoViReTs). We selected 22 LoViReTs matched with 22 controls ART suppressed for more than 3 years with fewer than 100 and more than 100 HIV-DNA copies/10 6 CD4 + T cells, respectively. We measured HIV reservoirs in blood and host genetic factors. Fourteen LoViReTs underwent leukapheresis to analyze replication-competent virus, and HIV-DNA in CD4 + T-cell subpopulations. Additionally, we measured HIV-DNA in rectum and/or lymph node biopsies from nine of them. We found that LoViReTs harbored not only lower levels of total HIV-DNA, but also significantly lower intact HIV-DNA, cell-associated HIV-RNA, and ultrasensitive viral load than controls. The proportion of intact versus total proviruses was similar in both groups. We found no differences in the percentage of host factors. In peripheral blood, 71% of LoViReTs had undetectable replication-competent virus. Minimum levels of total HIV-DNA were found in rectal and lymph node biopsies compared with HIV-infected individuals receiving ART. The main contributors to the reservoir were short-lived transitional memory and effector memory T cells (47% and 29%, respectively), indicating an altered distribution of the HIV reservoir in the peripheral T-cell subpopulations of LoViReTs. In conclusion, LoViReTs are characterized by low levels of viral reservoir in peripheral blood and secondary lymphoid tissues, which might be explained by an altered distribution of the proviral HIV-DNA towards more short-lived memory T cells. LoViReTs can be considered exceptional candidates for future interventions aimed at curing HIV

Neural Stem/Progenitor Cells from the Adult Human Spinal Cord Are Multipotent and Self-Renewing and Differentiate after Transplantation

Neural stem/progenitor cell (NSPC) transplantation is a promising therapy for spinal cord injury (SCI). However, little is known about NSPC from the adult human spinal cord as a donor source. We demonstrate for the first time that multipotent and self-renewing NSPC can be cultured, passaged and transplanted from the adult human spinal cord of organ transplant donors. Adult human spinal cord NSPC require an adherent substrate for selection and expansion in EGF (epidermal growth factor) and FGF2 (fibroblast growth factor) enriched medium. NSPC as an adherent monolayer can be passaged for at least 9 months and form neurospheres when plated in suspension culture. In EGF/FGF2 culture, NSPC proliferate and primarily express nestin and Sox2, and low levels of markers for differentiating cells. Leukemia inhibitory factor (LIF) promotes NSPC proliferation and significantly enhances GFAP expression in hypoxia. In differentiating conditions in the presence of serum, these NSPC show multipotentiality, expressing markers of neurons, astrocytes, and oligodendrocytes. Dibutyryl cyclic AMP (dbcAMP) significantly enhances neuronal differentiation. We transplanted the multipotent NSPC into SCI rats and show that the xenografts survive, are post-mitotic, and retain the capacity to differentiate into neurons and glia

Clonal Characterization of Rat Muscle Satellite Cells: Proliferation, Metabolism and Differentiation Define an Intrinsic Heterogeneity

Satellite cells (SCs) represent a distinct lineage of myogenic progenitors responsible for the postnatal growth, repair and maintenance of skeletal muscle. Distinguished on the basis of their unique position in mature skeletal muscle, SCs were considered unipotent stem cells with the ability of generating a unique specialized phenotype. Subsequently, it was demonstrated in mice that opposite differentiation towards osteogenic and adipogenic pathways was also possible. Even though the pool of SCs is accepted as the major, and possibly the only, source of myonuclei in postnatal muscle, it is likely that SCs are not all multipotent stem cells and evidences for diversities within the myogenic compartment have been described both in vitro and in vivo. Here, by isolating single fibers from rat flexor digitorum brevis (FDB) muscle we were able to identify and clonally characterize two main subpopulations of SCs: the low proliferative clones (LPC) present in major proportion (∼75%) and the high proliferative clones (HPC), present instead in minor amount (∼25%). LPC spontaneously generate myotubes whilst HPC differentiate into adipocytes even though they may skip the adipogenic program if co-cultured with LPC. LPC and HPC differ also for mitochondrial membrane potential (ΔΨm), ATP balance and Reactive Oxygen Species (ROS) generation underlying diversities in metabolism that precede differentiation. Notably, SCs heterogeneity is retained in vivo. SCs may therefore be comprised of two distinct, though not irreversibly committed, populations of cells distinguishable for prominent differences in basal biological features such as proliferation, metabolism and differentiation. By these means, novel insights on SCs heterogeneity are provided and evidences for biological readouts potentially relevant for diagnostic purposes described

TRIM5 Suppresses Cross-Species Transmission of a Primate Immunodeficiency Virus and Selects for Emergence of Resistant Variants in the New Species

Cross-species transmission of simian immunodeficiency virus from sooty mangabeys (SIVsm) into rhesus macaques, and subsequent emergence of pathogenic SIVmac, required adaptation to overcome restriction encoded by the macaque TRIM5 gene

The development and validation of a scoring tool to predict the operative duration of elective laparoscopic cholecystectomy

Background: The ability to accurately predict operative duration has the potential to optimise theatre efficiency and utilisation, thus reducing costs and increasing staff and patient satisfaction. With laparoscopic cholecystectomy being one of the most commonly performed procedures worldwide, a tool to predict operative duration could be extremely beneficial to healthcare organisations. Methods: Data collected from the CholeS study on patients undergoing cholecystectomy in UK and Irish hospitals between 04/2014 and 05/2014 were used to study operative duration. A multivariable binary logistic regression model was produced in order to identify significant independent predictors of long (> 90 min) operations. The resulting model was converted to a risk score, which was subsequently validated on second cohort of patients using ROC curves. Results: After exclusions, data were available for 7227 patients in the derivation (CholeS) cohort. The median operative duration was 60 min (interquartile range 45–85), with 17.7% of operations lasting longer than 90 min. Ten factors were found to be significant independent predictors of operative durations > 90 min, including ASA, age, previous surgical admissions, BMI, gallbladder wall thickness and CBD diameter. A risk score was then produced from these factors, and applied to a cohort of 2405 patients from a tertiary centre for external validation. This returned an area under the ROC curve of 0.708 (SE = 0.013, p  90 min increasing more than eightfold from 5.1 to 41.8% in the extremes of the score. Conclusion: The scoring tool produced in this study was found to be significantly predictive of long operative durations on validation in an external cohort. As such, the tool may have the potential to enable organisations to better organise theatre lists and deliver greater efficiencies in care

Selective transport of SC1 mRNA, encoding a putative extracellular matrix glycoprotein, during postnatal development of the rat cerebellum and retina

The selective transport of mRNA species into peripheral processes of cells is an important aspect of gene expression in the nervous system. In this study, we report the transport of SC1 mRNA into the distal processes of Bergmann glial (BG) cells at particular stages of development. SC1 is a putative anti-adhesive extracellular matrix (ECM) glycoprotein that is expressed not only in the developing central nervous system (CNS) but also in the adult brain. The intracellular distribution of SC1 mRNA was examined in two highly laminated neural structures, the cerebellum and retina, during postnatal development and in the adult rat. Our results indicate that SC1 mRNA expression is both spatially and temporally regulated. SC1 message was localized to BG cell bodies at postnatal day 5 (P5) and P10. However, by P15 through to the adult, SC1 mRNA was transported to distal processes of BG cells in the synapse-rich molecular layer (ML) of the cerebellum. In the developing rat retina, SC1 mRNA was expressed in specific neuronal populations by P10, however, transport of SC1 message to the dendrites of these retinal neurons was not detected during development or in the adult. These results indicate neural mechanisms which control the timing and cell type in which selective transport of SC1 mRNA is observed. The localization of SC1 mRNA to the distal processes of BG cells in the synapse-rich ML of the cerebellum could facilitate local control of SC1 protein synthesis which may play roles in synapse formation during development and in synaptic plasticity in the adult

Differential mRNA expression of the related extracellular matrix glycoproteins SC1 and SPARC in the rat embryonic nervous system and skeletal structure

SPARC is a multifunctional extracellular matrix glycoprotein that shares partial sequence homology with SC1. These extracellular matrix molecules are thought to play important roles in modulating cellular interactions. In vitro, SPARC has been shown to exhibit anti-adhesive activity. In the present investigation, in situ hybridization is used to compare the expression patterns of SC1 and SPARC mRNA in the rat embryo. Results show that SC1 and SPARC expression is spatially and temporally regulated. SC1 mRNA is strongly expressed in the embryonic brain and spinal cord, whereas SPARC mRNA is enriched in craniofacial cartilage and skeletal structures. This differential expression pattern in the rat embryo suggests that SC1 plays an important role in the developing nervous system, whereas SPARC participates primarily in events associated with skeletal development. However at embryonic day 17, SC1 and SPARC mRNA show parallel expression patterns in areas of the cerebellum undergoing cell migratory events

Expression of mRNA encoding extracellular matrix glycoproteins SPARC and SC1 is temporally and spatially regulated in the developing cochlea of the rat inner ear

SPARC is a multifunctional extracellular matrix (ECM) glycoprotein that shares partial sequence homology with SC1/hevin. These ECM molecules exhibit calcium-binding properties and modulate cellular interactions. This study examines the expression of SC1 and SPARC mRNA in the developing cochlea of the rat inner ear prior to and after the onset of hearing. At all ages examined, SC1 mRNA is highly expressed in neurons of the spiral ganglion. In contrast, SPARC transcripts are not detected in the spiral ganglion but are enriched in the temporal bone and cartilaginous otic capsule surrounding the cochlea. Both SC1 and SPARC mRNA are expressed in connective tissue elements involved in maintaining ionic homeostasis of cochlear fluids. SC1 mRNA is localized to type III fibrocytes of the spiral ligament (slg) and marginal cells of the stria vascularis, while SPARC mRNA is apparent in the spiral limbus and type I fibrocytes of the slg. At postnatal day 10, SPARC mRNA shows a dramatic change in expression. High levels of SPARC transcripts are induced in Deiters cells (dc) of the organ of Corti. Interestingly, this induction of SPARC mRNA correlates with the onset of hearing. This suggests that SPARC may play a role in calcium regulation in dc when functional maturation of the cochlea is attained and rapid changes in calcium levels are required